Roles Of NPY1R And OLFM1 In The Progress Of Estrogen-Receptor-?-Positive Breast Cancer

Objective:To study the function of NPY1R and OLFM1 in ER?-positive breast cancer and to explore the relationship between the expression of NPY1R and OLFM1 and clinicopathological features and survival of patients with breast cancer.Methods:By using publicly available gene expression profiling datasets,GOBO,we observed the expression of NPY1R or OLFM1 in different molecular subtypes of breast cancer cell lines and patients.Also we compared the association between NPY1R or OLFM1 and survival of breast cancer patients with GOBO.Moreover,we choose MCF7 cells to study the functions.E2 was used to enhance ER?signaling pathway to confirm if ER?can regulate the expression of NPY1R or OLFM1.After silenced NPY1R and OLFM1 expression through RNA interference system,we use WST-1?Roche?,Matrigel Invasion Chambers,Cell Death Detection ELISA^PLUS Kit and Western Blot to detect cell proliferation,invasion and apoptosis.In addition,IBM SPSS 24.0 was used to analysis the relationship between gene expression and clinicopathological factors,also the prognostic significance of gene expression and all clinical covariates after identify the expression of NPY1R and OLFM1 by using immunohistochemical staining.Results:1.NPY1R expression in ER?-Positive Breast CancerUsing GOBO online database we observed that lower expression of NPY1R in triple-negative and HER2 breast cancer cell lines,but higher expression in some of the ER?-positive breast cancer cell lines?P<0.05?.Patients with higher NPY1R expression had worse DMFS among all breast cancer patients?P<0.05?.In ER?-positive breast cancer patients,the expression of NPY1R was much higher?P<0.01?,and associated with poor prognosis?P<0.05?.Based on MCF7 cell model,we found E2 can upregulate the expression of NPY1R at mRNA and protein level.In addition,knockdown of NPY1R inhibited cell proliferation and invasion,promote apoptosis.To explore the expression of NPY1R in breast carcinogenesis,immunohistochemistry staining was carried out in breast cancer tissues and paracancerous tissues.We found there was significant difference between the expression level of NPY1R in this two kind of tissues?P<0.05?.Furthermore,we found NPY1R expression was significantly associated with histologic grade?P<0.05?,but the relationship among the expression level of NPY1R and the clinicopathological factors?Age,Tumor location,Lymph node status,T stage,AJCC stage?,molecular subtype factors?ER?,PR,HER2?,prognostic factors?S100,P53?,the correlation factors of malignant degree?SMA,CK7,BCL-2,HCK?and metastatic correlation factors?E-cadherin?had no significant difference?P>0.05?.Univariate analysis showed that histologic grade,lymph node status,AJCC stage,ER?status,P53 status and NPY1R expression status were significant associated with overall survival.Muitivariate COX analysis revealed that NPY1R expression status,ER?status and P53 status were confirmed as significant prognostic factors?P<0.05?.Survival analysis results showed higher NPY1R expression was associated with poor outcome,especially in lymph node positive status in breast cancer?P<0.05?.However,no significant differences were observed in patients whose age were over 50,ER?-positive subtype,stage 2-3 subgroup or in histologic grade 2-3 with higer NPY1R expression?P>0.05?.2.OLFM1 expression in ER?-Positive Breast CancerWe found that OLFM1 was overexpression in ER?-positive breast cancer cell lines?P<0.05?.And the expression of OLFM1 was upregulated after treated with E2.Moreover,silencing of OLFM1 reduced the cell proliferation and invasion ability,and enhanced apoptosis ability.In addition,we study the relationship between OLFM1 expression and clinical parameters.Immunohistochemistry staining results show that there has no significant difference between the expression level of OLFM1 in breast cancer tissues and paracancerous tissues?P>0.05?.However,GOBO online database showed the expression of OLFM1 was higher in ER?-positive breast cancer patients?P<0.01?.After analysed the relationship among the expression level of OLFM1 and the clinicopathological factors?Age,Tumor location,Histologic grade,Lymph node status,T stage,AJCC stage?,molecular subtype factors?ER?,PR,HER2?,prognostic factors?S100,P53?,the correlation factors of malignant degree?SMA,CK7,BCL-2,HCK?and metastatic correlation factors?E-cadherin?,we found OLFM1 expression was significantly associated with histologic grade and E-cadherin status,but not in the others?P<0.05?.Univariate analysis showed that histologic grade,lymph node status,AJCC stage,ER?status and P53 status were significant associated with overall survival.Muitivariate COX analysis revealed that OLFM1 expression status,ER?status and P53 status were confirmed as significant prognostic factors?P<0.05?.GOBO online database showed patients with high OLFM1 expression had better DMFS than patients with low OLFM1 expression?P<0.05?.Survival analysis of different breast cancer subgroup results showed patients whose age were over 50,ER?-positive subtype or in stage 2-3 subgroup with higher OLFM1 expression had worse OS than patients with lower OLFM1 expression?P<0.05?.Patients in histologic grade 2-3 or lymph node positive status with higher OLFM1 expression had worse OS than patients with lower OLFM1 expression,but no significant differences were found?P>0.05?.Conclusion:1.ER?upregulates expression of NPY1R or OLFM1 at mRNA and protein level.2.Knockdown of NPY1R or OLFM1 inhibite cell proliferation and invasion,promote apoptosis.3.NPY1R expression status,OLFM1 expression status,ER?status and P53 status were confirmed as significant prognostic factors4.Breast cancer patients or patients in lymph node positive status with higher NPY1R expression had worse OS than patients with lower NPY1R expression.5.ER?-positive subtype patients,patients whose age was over 50 or patients in stage 2-3 subgroup with higher OLFM1 expression had worse OS than patients with lower OLFM1 expression.